Ethanol treatment enhances drought stress avoidance in cassava (Manihot esculenta Crantz)

Plant Molecular Biology - External application of ethanol enhances tolerance to high salinity, drought, and heat stress in various plant species. However, the effects of ethanol application on...     

Accumulation of Small Heat-Shock Protein Homologs in the Endoplasmic Reticulum of Cortical Parenchyma Cells in Mulberry in Association with Seasonal Cold Acclimation

Cortical parenchyma cells of mulberry (Morus bombycis Koidz.) trees acquire extremely high freezing tolerance in winter as a result of seasonal cold acclimation. The amount of total proteins in endoplasmic reticulum (ER)-enriched fractions isolated from these cells increased in parallel with the process of cold acclimation. Protein compositions in the ER-enriched fraction also changed seasonally, with a prominent accumulation of 20-kD (WAP20) and 27-kD (WAP27) proteins in winter. The N-terminal amino acid sequence of WAP20 exhibited homology to ER-localized small heat-shock proteins (smHSPs), whereas that of WAP27 did not exhibit homology to any known proteins. Like other smHSPs, WAP20 formed a complex of high molecular mass in native-polyacrylamide gel electrophoresis. Furthermore, not only WAP20 but also 21-kD proteins reacted with antibodies against WAP20. Fractionation of the crude microsomes by isopycnic sucrose-gradient centrifugation revealed that both WAP27 and WAP20 were distributed on a density corresponding to the fractions with higher activity of ER marker enzyme, suggesting localization of these proteins in the ER. When ER-enriched fractions were treated with trypsin in the absence of detergent, WAP20 and WAP27 were undigested, suggesting localization of these proteins inside the ER vesicle. The accumulation of a large quantity of smHSPs in the ER in winter as a result of seasonal cold acclimation indicates that these proteins may play a significant role in the acquisition of freezing tolerance in cortical parenchyma cells of mulberry trees.     

Monoclonal antibody ECCD-1 inhibits intercellular communication in teratocarcinoma PCC3 cells

The monoclonal antibody ECCD-1 recognizing a certain class of cell surface proteins inhibits the Ca²⁺-dependent cell-to-cell adhesion in teratocarcinoma stem cells. In this paper, we studied the effect of ECCD-1 on cell-to-cell communication in PCC3 cells by measuring the transfer of lucifer yellow between cells. To this aim, PCC3 cells were cultured in the presence of ECCD-1 for various periods, and then the fluorescent dye was injected into a cell located in the center of cell colonies, followed by counting number of cells to which the dye was transferred. The results showed that ECCD-1 inhibits the dye transfer between cells, suggesting that the Ca²⁺-dependent cell-to-cell adhesion system (CDS) is essential for the functions of gap junction.     

Distinct Cellular Expression of Calcineurin Aα and Aβ in Rat Brain

Specific polyclonal antibodies that distinguish the two distinct isoforms of the catalytic subunit of calmodulin-dependent protein phosphatase, calcineurin A alpha and A beta, were prepared, and the distribution of calcineurin A alpha and A beta in rat brain was studied using immunochemical and immunocytochemical techniques. Immunochemical measurement revealed that the regional distributions of the two isoforms differed and that A alpha was more abundant than A beta in the rat brain. The subcellular distribution patterns of both isoforms were similar. Both isoforms were highly enriched in cytosolic fractions, including the synaptosomal cytosol. Immunocytochemical analysis revealed that both A alpha and A beta immunoreactivities differed in regional and cellular localizations. These different patterns of expression suggest that the two isoforms of calcineurin A may each have specific functions in modulating neuronal activity in particular cell types.     

IMMUNOLOCALIZATION OF CENTRIN DURING FERTILIZATION AND THE FIRST CELL CYCLE IN FUCUS DISTICHUS AND PELVETIA COMPRESSA (FUCALES,PHAEOPHYCEAE)1

Antibodies that recognize the centrosome-associated protein centrin were used to characterize centrosomal origin and positioning during fertilization and the first cell cycle in Fucus distichus subsp. evanescens (C. Agardh) Powell and Pelvetia compressa (J. Agardh) De Toni. Centrin was identified in sperm, eggs, and zygotes on protein blots, indicating the protein is present in both gametes. Using immunofluorescence microscopy, centrin was found in discrete foci in sperm. In contrast, eggs lack centrosomes and centrin was not detectable by immunofluorescence, indicating that centrin was probably dispersed in the cytoplasm. Two foci of centrin were present on the nuclear envelope of zygotes, but microtubules remained dispersed over the zygotic nucleus. Centrin foci separated over the nuclear envelope as the first cell cycle progressed. Microtubules became concentrated at the centrin foci to form centrosomes that gave rise to the spindle poles at mitosis.     

A novel method for determination of α1,6fucosyltransferase activity using a reducing oligosaccharide from egg yolk as a specific acceptor

A new method for determination of 1,6fucosyltransferase activity has been described. Recently, the disialyl-biantennary undecasaccharide was prepared in high yield from egg yolk [(1996), Carbohydr Lett 2: 137–42]. By treatment of this oligosaccharide with neuraminidase and -galactosidase, we readily obtained an asialo-agalacto-biantennary heptasaccharide (GlcNAc 1,2Man1,6[GlcNAc1,2Man1,3]Man1,4GlcNAc1,4GlcNAc). Using this asialo-agalacto-oligosaccharide as an acceptor, fucosyltransferases from human plasma and extracts of various human hepatoma cell lines were assayed in the presence of GDP-[3H]fucose. The reaction mixture was applied to a column of GlcNAc-binding, Psathyrella velutina lectin coupled gel. All the fucosylated acceptor were bound to the column which was eluted with 50 mM GlcNAc. Structural analyses revealed that only the innermost GlcNAc residue of the acceptor was fucosylated through an 1,6-linkage, and the oligosaccharide prepared could be used as a specific acceptor for 1,6fucosyltransferase. The present method was used to screen plasma 1,6fucosyltransferase in several patient groups, and significantly elevated activities were found in samples from patients with liver diseases, including chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma.     

Complete Nucleotide Sequences of 93-kb and 3.3-kb Plasmids of an Enterohemorrhagic Escherichia coli O157:H7 Derived from Sakai Outbreak

Enterohemorrhagic Escherichia coli (EHEC) O157:H7, derived froman outbreak in Sakai city, Japan in 1996, possesses two kindsof plasmids: a 93-kb plasmid termed pO157, found in clinicalEHEC isolates world-wide and a 3.3-kb plasmid termed pOSAK1,prevalent in EHEC strains isolated in Japan. Complete nucleotidesequences of both plasmids have been determined, and the putativefunctions of the encoded proteins and the cis-acting DNA sequenceshave been analyzed. pO157 shares strikingly similar genes andDNA sequences with F-factor and the transmissible drug-resistantplasmid R100 for DNA replication, copy number control, plasmidsegregation, conjugative functions and stable maintenance inthe host, although it is defective in DNA transfer by conjugationdue to the truncation and deletion of the required genes andDNA sequences. In addition, it encodes several proteins implicatedin EHEC pathogenicity such as an EHEC hemolysin (HlyA), a catalase-peroxidase(KatP), a serine protease (EspP) and type II secretion system.pOSAK1 possesses a ColE1-like replication system, and the DNAsequence is extremely similar to that of a drug-resistant plasmid,NTP16, derived from Salmonella typhimurium except that it lacksdrug resistance transposons.     

KARYOGAMY BLOCK BY HEAT STRESS IN THE FERTILIZATION OF BROWN ALGAE

Karyogamy was inhibited by heat stress in zygotes of Scytosiphon lomentaria (Lyngbye) Link (isogamy), Cutleria cylindrica Okamura (anisogamy), and Fucus distichus subsp. evanescens (C. Agardh) Powell (oogamy). Although high temperatures did not inhibit migration of the male and female nuclei, nuclear envelope fusion was blocked. The ultrastructural stage at which karyogamy was inhibited varied among these species. In S. lomentaria, the outer membranes fused with each other, but the inner membranes did not fuse. Partial fusion of the nuclear envelope occurred in C. cylindrica. In F. distichus, the block of karyogamy at high temperature was incomplete, and nuclear fusion proceeded gradually. The block to karyogamy in S. lomentaria zygotes was reversible, and karyogamy proceeded when zygotes were transferred from 22° to 14° C. Experiments using inhibitors suggested that proteins that might be formed de novo after fertilization do not participate in karyogamy or its inhibition at either 14° or 22° C.